A novel method to determine the diffusional water permeability of oocyte plasma membranes.
Measurements of the cell membrane diffusional water permeability (Pd) are important to characterize water passage across water channels and across the lipid bilayer component of the membrane. Existing methods for those measurements are involved; however, we report here a simple procedure to estimate Pd in Xenopus laevis oocytes and similar large cells. Due to the different densities of H2O and D2O (heavy water), an oocyte transferred from normal medium to a D2O-based medium floats initially, but subsequently sinks when a certain amount of the water originally in them is replaced by the D2O that diffuses in. We describe how the 'flotation time' (time that oocytes float in a heavy water solution before they start sinking) yields the Pd of the plasma membrane. Determination of Pd by this procedure and by the rate of tritiated water (T2O) efflux give for Pd results which are very close: 2.2 +/- 0.2 (n = 8) and 2.0 +/- 0.1 (n = 6) microns/s, respectively (T = 10 degrees C). Furthermore, our method detects the increase in Pd elicited in oocytes by either expression of water channel proteins, or by treating them with the pore-forming antibiotic amphotericin B. This method appears useful to gauge the expression and function of pore-forming, water-permeable membrane proteins.
Iserovich
P
P
Department of Physiology, College of Physicians and Surgeons, Columbia University, New York, NY 10032, USA.
Kuang
K
K
Chun
T
T
Fischbarg
J
J
D000666:Amphotericin B / Q000494:pharmacology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002463:Cell Membrane Permeability; D003903:Deuterium / Q000493:pharmacokinetics*; D007473:Ion Channels / Q000235:genetics; D008845:Microinjections; D009865:Oocytes / Q000737:chemistry / Q000378:metabolism; D009995:Osmosis; D012333:RNA, Messenger / Q000494:pharmacology; D014316:Tritium / Q000493:pharmacokinetics*; D014867:Water / Q000378:metabolism*; D014982:Xenopus laevis
D000900:Anti-Bacterial Agents; D007473:Ion Channels; D012333:RNA, Messenger; D014867:Water; D014316:Tritium; D000666:Amphotericin B; D003903:Deuterium